New domain of poly-gamma-glutamate synthase PgsB
Ontology highlight
ABSTRACT:
ORGANISM(S): bacillus
SUBMITTER: zhanglei
PROVIDER: S-BSST430 | biostudies-other |
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Poly-γ-glutamate (PGA), a novel polyamide material with industrial applications, possesses a nylon-like backbone, is structurally similar to polyacrylic acid, is biodegradable and is safe for human consumption. PGA is frequently found in the mucilage of natto, a Japanese traditional fermented food. To date, three different types of PGA, namely a homo polymer of D-glutamate (D-PGA), a homo polymer of L-glutamate (L-PGA), and a random copolymer consisting of D- and L-glutamate (DL-PGA), are known. This review will detail the occurrence and physiology of PGA. The proposed reaction mechanism of PGA synthesis including its localization and the structure of the involved enzyme, PGA synthetase, are described. The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance. Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed. Finally, some techniques for PGA and its derivatives in advanced material technology are presented.
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Project description:Some Bacillus subtilis strains, including natto (fermented soybeans) starter strains, produce a capsular polypeptide of glutamate with a gamma-linkage, called poly-gamma-glutamate (gamma-PGA). We identified and purified a monomeric 25-kDa degradation enzyme for gamma-PGA (designated gamma-PGA hydrolase, PghP) from bacteriophage PhiNIT1 in B. subtilis host cells. The monomeric PghP internally hydrolyzed gamma-PGA to oligopeptides, which were then specifically converted to tri-, tetra-, and penta-gamma-glutamates. Monoiodoacetate and EDTA both inhibited the PghP activity, but Zn(2+) or Mn(2+) ions fully restored the enzyme activity inhibited by the chelator, suggesting that a cysteine residue(s) and these metal ions participate in the catalytic mechanism of the enzyme. The corresponding pghP gene was cloned and sequenced from the phage genome. The deduced PghP sequence (208 amino acids) with a calculated M(r) of 22,939 was not significantly similar to any known enzyme. Thus, PghP is a novel gamma-glutamyl hydrolase. Whereas phage PhiNIT1 proliferated in B. subtilis cells encapsulated with gamma-PGA, phage BS5 lacking PghP did not survive well on such cells. Moreover, all nine phages that contaminated natto during fermentation produced PghP, supporting the notion that PghP is important in the infection of natto starters that produce gamma-PGA. Analogous to polysaccharide capsules, gamma-PGA appears to serve as a physical barrier to phage absorption. Phages break down the gamma-PGA barrier via PghP so that phage progenies can easily establish infection in encapsulated cells.
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Project description:Residual dipolar couplings (RDCs) offer additional information for structure elucidation by NMR spectroscopy. They are measured in anisotropic media, such as lyotropic liquid crystalline phases of polypeptides. Today, some suitable polypeptides are known. Nevertheless, structural influences of these polypeptides on the alignment properties are not really understood. Thus, which influence a chiral side chain has on enantiodiscrimination and whether we can improve the enantiodifferentiation significantly by adding an additional chiral center in the side chain are questions of interest. Therefore, new diastereomeric polypeptide-based alignment media with an additional chiral center in the side chain derived from perillyl alcohol were synthesized and their properties were investigated (secondary structure, liquid crystallinity, etc.). The enantiomers of isopinocampheol and β-pinene were used as model analytes for the study of enantiodiscrimination. Additionally, the usage of 1 H-1 H-RDCs to improve the alignment tensor quality is demonstrated.
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Project description:For many years, silver nanoparticles, as with other antibacterial nanoparticles, have been extensively used in manufactured products. However, their fate in the environment is unclear and raises questions. We studied the fate of silver nanoparticles in the presence of bacteria under growth conditions that are similar to those found naturally in the environment (that is, bacteria in a stationary phase with low nutrient concentrations). We demonstrated that the viability and the metabolism of a gram-positive bacteria, Bacillus subtilis, exposed during the stationary phase is unaffected by 1 mg/L of silver nanoparticles. These results can be partly explained by a physical interaction of the poly-gamma-glutamate (PGA) secreted by Bacillus subtilis with the silver nanoparticles. The coating of the silver nanoparticles by the secreted PGA likely results in a loss of the bioavailability of nanoparticles and, consequently, a decrease of their biocidal effect.
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Project description:The identification of new strategies to fight bacterial infections in view of the spread of multiple resistance to antibiotics has become mandatory. It has been demonstrated that several bacteria develop poly-γ-glutamic acid (γ-PGA) capsules as a protection from external insults and/or host defence systems. Among the pathogens that shield themselves in these capsules are Bacillus anthracis, Francisella tularensis and several Staphylococcus strains. These are important pathogens with a profound influence on human health. The recently characterised γ-PGA hydrolases, which can dismantle the γ-PGA-capsules, are an attractive new direction that can offer real hope for the development of alternatives to antibiotics, particularly in cases of multidrug resistant bacteria. We have characterised in detail the cleaving mechanism and stereospecificity of the enzyme PghL (previously named YndL) from Bacillus subtilis encoded by a gene of phagic origin and dramatically efficient in degrading the long polymeric chains of γ-PGA. We used X-ray crystallography to solve the three-dimensional structures of the enzyme in its zinc-free, zinc-bound and complexed forms. The protein crystallised with a γ-PGA hexapeptide substrate and thus reveals details of the interaction which could explain the stereospecificity observed and give hints on the catalytic mechanism of this class of hydrolytic enzymes.
| S-EPMC6506827 | biostudies-literature
Project description:Particular Bacillus subtilis strains produce a capsular polypeptide poly-gamma-glutamate (gamma-PGA) that functions as a physical barrier against bacteriophage infection. Bacteriophage PhiNIT1 can infect B. subtilis and produces a novel gamma-PGA hydrolase PghP. PghP was overexpressed, purified and crystallized by the sitting-drop vapour-diffusion method. The crystals diffracted to a resolution of 2.4 A using a synchrotron X-ray source and were found to belong to space group P3(1)21 or P3(2)21.
| S-EPMC2795599 | biostudies-literature
Project description:Poly-γ-glutamate (PGA) possesses a nylon-like backbone and polyacrylate-like carboxyl groups, and shows an extraordinary solubility in water. In this study, the effective synthesis and structural analysis of some water-insoluble PGA ion-complexes (PGAICs) using cationic surfactants, hexadecylpyridinium (HDP), dodecylpyridinium, benzalkonium and benzetonium, were examined. We demonstrated their spontaneous coating performance to the surfaces of different materials (i.e., plastics, metals, and ceramics) as potent anti-staphylococcal and anti-Candida agents. The tests against Staphylococcus aureus revealed that, regardless of a variety of materials, PGAICs maintained surface antimicrobial activity, even after the water-soaking treatment, whereas those against Candida albicans indicated that, among PGAICs, PGA/HDP complex is most useful as an anti-fungal agent because of its coating stability. Moreover, the log reduction values against Influenza A and B viruses of PGA/HDP-coated surfaces were estimated to be 5.4 and 3.2, respectively, suggesting that it can be dramatically suppressed the infection of influenza. This is to our knowledge the first observation of PGA-based antiviral coatings.
| S-EPMC4632766 | biostudies-literature
Project description:We are now entering a new age of intelligent material development using fine, sustainable polymers from extremophiles. Herein we present an innovative (but simple) means of transforming archaeal poly-γ-glutamate (PGA) into extremely durable polyionic complexes with potent antimicrobial performance. This new supra-polymer material (called PGA/DEQ) was subjected to nuclear magnetic resonance and X-ray diffraction spectroscopies to characterize in structural chemistry. Calorimetric measurements revealed its peculiar thermal properties; to the best of our knowledge, it is one of the most heat-resistant biopolymer-based polyionic complexes developed to date. PGA/DEQ is particularly useful in applications where surface functionalization is important, e.g., antimicrobial coatings. The spontaneously assembled PGA/DEQ coatings (without any additional treatments) were remarkably resistant to certain organic solvents (including chloroform), even at high salt concentrations (theoretically greater than those found in sea water), and various pH values. However, the pH-response tests also implied that the PGA/DEQ coatings could be removed only when concentrated citrate di-salts were used, whereas most crosslinked polymer composites (e.g., thermoset matrices) are difficult to recycle and treat downstream. We also discuss PGA/DEQ-immobilized surfaces that exhibit enigmatic microbicidal mechanisms.
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Project description:Bacillus subtilis swims in liquid media and swarms over solid surfaces, and it encodes two sets of flagellar stator homologs. Here, we show that B. subtilis requires only the MotA/MotB stator during swarming motility and that the residues required for stator force generation are highly conserved from the Proteobacteria to the Firmicutes. We further find that mutants that abolish stator function also result in an overproduction of the extracellular polymer poly-γ-glutamate (PGA) to confer a mucoid colony phenotype. PGA overproduction appeared to be the result of an increase in the expression of the pgs operon that encodes genes for PGA synthesis. Transposon mutagenesis was conducted to identify insertions that abolished colony mucoidy and disruptions in known transcriptional regulators of PGA synthesis (Com and Deg two-component systems) as well as mutants defective in transcription-coupled DNA repair (Mfd)-reduced expression of the pgs operon. A final class of insertions disrupted proteins involved in the assembly of the flagellar filament (FliD, FliT, and FlgL), and these mutants did not reduce expression of the pgs operon, suggesting a second mechanism of PGA control.
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Project description:Previously, we reported that the oral administration of high molecular mass poly-gamma-glutamate (gamma-PGA) induced antitumor immunity but the mechanism underlying this antitumor activity was not understood. In the present study, we found that application of high molecular mass gamma-PGA induced secretion of tumor necrosis factor (TNF)-alpha from the bone-marrow-derived macrophages of wild type (C57BL/6 and C3H/HeN) and Toll-like receptor 2 knockout (TLR2(-/-)) mice, but not those of myeloid differentiation factor 88 knockout (MyD88(-/-)) and TLR4-defective mice (C3H/HeJ). Production of interferon (IFN)-gamma-inducible protein 10 (IP-10) in response to treatment with gamma-PGA was almost abolished in C3H/HeJ mice. In contrast to LPS, gamma-PGA induced productions of TNF-alpha and IP-10 could not be blocked by polymyxin B. Furthermore, gamma-PGA-induced interleukin-12 production was also impaired in immature dendritic cells (iDCs) from MyD88(-/-) and C3H/HeJ mice. Downregulation of MyD88 and TLR4 expression using small interfering RNA (siRNA) significantly inhibited gamma-PGA-induced TNF-alpha secretion from the RAW264.7 cells. Gamma-PGA-mediated intracellular signaling was markedly inhibited in C3H/HeJ cells. The antitumor effect of gamma-PGA was completely abrogated in C3H/HeJ mice compared with control mice (C3H/HeN) but significant antitumor effect was generated by the intratumoral administration of C3H/HeN mice-derived iDCs followed by 2,000 kDa gamma-PGA in C3H/HeJ. These findings strongly suggest that the antitumor activity of gamma-PGA is mediated by TLR4.
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